Tumor formation is often characterized by the uncoupling of the normally opposing processes of cellular proliferation and differentiation (Bishop, Cell 64:235-248 (1991); Hunter, Cell 64:249-270 (1991); and Sawyers et al., Cell 64:337-350 (1991)). The mechanisms that disrupt their normal coupling are believed to involve the overexpression or inappropriate expression of hematopoietic growth factors, growth factor receptors, and/or certain proto-oncogenes. The overexpression of the proto-oncogene c-myc has been detected in tumors, such as breast, stomach, lung, and colon carcinomas, neuroblastomas, and glioblastomas. Also, overexpression of c-myc induces neoplastic transformation of hematopoietic cells. For example, constitutive expression of the c-myc gene inhibits induced differentiation in erythroleukemia cells and in monocytic cells (Coppola and Cole, Nature 320:760-763 (1986)); Dmitrovsky et al. , Nature 322:748-750 (1986); Larsson et al., Proc. Natl. Acad. Sci. USA 85:2638-2642 (1988); Prochownik and Kukowska, Nature 322:848-850 (1986)). Furthermore, antisense oligonucleotides to the c-myc coding region prevent accumulation of c-Myc protein. This results in maturation of the target hematopoietic cells (Holt et al., Mol. Cell. Biol. 8:963-973 (1988)). These studies support the hypothesis that the level of c-myc messenger RNA (mRNA) and protein sets the balance between proliferation and differentiation. Tuning c-myc to low levels allows for differentiation, while high levels favor proliferation. The deregulation of this balance appears to be one genetic event which can ultimately lead to the uncontrolled cell proliferation that is characteristic of the neoplastic phenotype.
The c-myc gene is not unique in terms of having an essential role in cellular growth processes. It has been known for decades that specific and timely changes in the expression of multiple genes are required for proper embryonic development and cell maturation (Davidson, Gene Activity in Early Development, 3rd ed., Academic Press, Orlando, Fla. (1986)). However, studies have focused on c-myc, because it seems to be regulated not only at the levels of transcription, attenuation, nuclear processing, and translation, but also at the level of mRNA turnover (Marcu et al., Ann. Rev. Biochem. 61:809-860 (1992)). Indeed, direct half-life measurements indicated that c-myc mRNA has a half-life of 15-40 minutes (Dani et al., Proc Natl. Acad. Sci., USA 81:7046-7050 (1984); Dani et al., Proc. Natl. Acad. Sci., USA 82:4896-4899 (1985) and Piechaczyk et al., Cell 42:597-598 (1985)). These and other studies (Marcu et al., Ibid.) demonstrated that the control of c-myc mRNA turnover might be an important means of regulating both the level and timing of c-myc expression.
Many proto-oncogene mRNAs are very unstable. The rapid turnover of c-myc mRNA is controlled by sequences in the 3'-untranslated region (3'UTR) or by coding region sequences (Laird-Offringa, BioEssays 14:119-124 (1992) and Schiavi et al., Biochim. Biophy. Acta 1114:95-106 (1992)). A common feature in the labile mRNAs of proto-oncogenes, such as c-myc, c-fos and of the cytokine GM-CSF, is the presence of an AU-rich element (ARE) in the 3'UTR which is one cis-acting element responsible for their rapid degradation (Atwater et al, Annu. Rev. Genet. 24:519-541 (1990); Peltz et al., Crit. Rev. Euk. Gene Expression 1:99-126 (1991) and Schiavi et al., Biochim. Biophy. Acta 1114: 95-106 (1992)).
Functionally, the ARE appears to mediate sequentially rapid deadenylation which is followed by cleavage of the body of the mRNA (Brewer and Ross, Mol. Cell. Biol. 8:1697-1708 (1988); Shyu et al., Genes Dev. 5:221-231 (1991); Wilson and Treisman, Nature 336:396-399 (1989)). Several groups have identified ARE-binding proteins that might mediate the degradation of these mRNAs (Bickel et al., Proc. Natl., Acad. Sci. USA 89:10001-10005 (1992); Bohjanen et al., Mol. Cell. Biol. 11:3288-3295 (1991); Bohjanen et al., J. Biol. Chem. 267:6302-6309 (1992); Brewer, Mol. Cell. Biol. 11:2460-2466 (1991); Malter, Science 246:664-666 (1989); Myer et al., Proc. Natl. Acad. Sci. USA 89:1296-1300 (1992); Vakalopoulou et al. , Mol. Cell. Biol. 11:3355-3364 (1991); You et al., Mol. Cell. Biol. 12:2931-2940 (1992)). Recently, Hamilton et al., J. Biol. Chem. 268:8881-8887 (1993) identified two of these ARE-binding proteins as hnRNP A1 and C.
These ARE-binding proteins are capable of limiting the expression of proto-oncogenes whose overexpression activates the growth of cells found in malignant tumors. Therefore, the genes responsible for the expression of these ARE-binding proteins represent a new mechanism of tumor suppression, degrading proto-oncogene mRNAs to limit their expression. Previously, tumor suppressor genes were known only to encode proteins controlling transcription and translation. Only a limited number of tumor suppressor genes have been identified. To expand the number of therapeutic targets and diagnostic tools for cancer, additional tumor suppressor genes need to be identified.